Project Summary/Abstract The Drosophila Dscam gene encodes a vast family of immunoglobulin (Ig) superfamily proteins and plays a crucial role in regulating the formation of precisely organized neural circuits during development. Alternative splicing generates more than 38,016 different isoforms, including 19,008 ectodomains tethered to the membrane by one of two alternative transmembrane segments. These ectodomains share the same domain structure, but differ in amino acid sequence within three variable domains, Ig2, Ig3 and Ig7. Isoforms sharing the same combination of variable domains (i.e. they match at each variable domain) bind to each other, but they do not bind to other isoforms. Genetic analysis has revealed that Dscam plays a crucial role in a phenomenon called self-avoidance. Each neuron expresses a unique combination of isoforms and this allows its neurites to distinguish between self and non-self. Self-recognition, namely isoform-specific homophilic binding between neurites of the same cell, promotes repulsion between these processes. Self-avoidance plays an important role in the segregation of axon branches and the elaboration of dendritic fields and, thereby, contributes to the assembly of neural circuits. In this proposal, we will address three questions: 1. Is homophilic recognition crucial for all of Dscam's functions? To address this question we will generate knock-in mutants of Dscam which have lost the ability to engage in homophilic binding; 2. Are specific isoforms of Dscam expressed in distinct populations of neurons? While Dscam clearly plays a role in self-avoidance, it remains possible that different populations of neurons may use specific isoforms of Dscam to recognize other neurons. This aim is directed towards identifying such cells; and 3. Are their requirements in specific neurons for different alternative Ig domains? To address this question we will generate separate lines of flies with a single Ig2 domain, a single Ig3 domain or a single Ig7 domain. While Dscam diversity is not observed in mammals, Dscam is expressed in a cell-type specific fashion and appears to play an analogous function in self-avoidance in mouse amacrine cells. As the dosage of human Dscam has been implicated in Down Syndrome the studies described here may provide insight into the etiology of mental retardation associated with this syndrome.